The imbalance of reactive oxygen species (ROS) and reactive nitrogen species (RNS) turnover contributes to the progression of neurodegeneration. In the eye, low/mild ROS levels in cells/tissues could contribute to maintaining cellular homeostasis, signaling, and survival. In contrast, excessive ROS induces significant neuronal oxidative damage ^[1]. Therefore, balancing redox homeostasis contributes strongly to maintaining a proper level of ROS and RNS and, thus, healthy conditions. Mitochondria, essential in providing energy and other key cell functions, are a significant source of reactive species that can lead to the progression of pathologies, including those related to the visual system ^[2][3]. Still, ROS are crucial for communication between the mitochondria and the nucleus, although their overproduction could impair mitochondria, damaging mitochondrial DNA (mtDNA), proteins, lipids, and membranes. The resulting dysfunctional mitochondria could induce cellular apoptosis and neurodegeneration. Therefore, preserving proper mitochondrial homeostasis is essential for cells, including retinal neurons.

The retina is a part of the central nervous system, with a high metabolism and energy demands mainly satisfied by oxidative and glycolytic metabolism, resulting in ROS/RNS production and, possibly, accumulation. Within the retinal neurons, photoreceptors primarily require a high amount of energy. Indeed, photoreceptors are not only the first step in phototransduction but also represent a source of metabolic intermediates for the surrounding cells, being in close contact with the retinal pigment epithelium (RPE), which in turn is linked to the choroidal vasculature ^[4]. RPE supplies photoreceptors with plenty of glucose, which is crucial for the retinal neuron’s homeostasis and energy needs. Also, inside photoreceptors, mitochondria produce ATP using the fatty acids and ketone bodies that RPE supplies. In physiological conditions, different cells enact strategies to hamper metabolic impairments inside the retina ^[1]. Müller cells, one of the three types of retinal glial cells, fulfill the delicate role of supervising neurons’ necessities, including metabolic conditions. Activated Müller glia contribute to counterbalancing metabolic impairments. Still, its prolonged activation can lead to harmful conditions, such as inflammation and vascular defects. Also, retinal ganglion cells (RGCs), the inner-most retinal neurons, maintain homeostasis in the retina and regulate the retinal blood flow by secreting angiogenic factors, thus supervising the supply of nutrients to the entire retina.

Due to their central role in sending action potentials to the brain, RGCs consume high energy for their metabolism and are particularly susceptible to mitochondrial dysfunction. RGCs mitochondrial impairment hampered ATP supply and was related to severe pathologies hitting the optic nerve and eventually causing glaucoma ^[5], or the entire neural retina, such as in DR. Not less noteworthy, mitochondrial dysfunction causes oxidative stress in RGCs, which contributes to neurodegeneration ^[6][7]. Under acute stress, RGCs can efficiently end damaged mitochondria, activating biogenesis to maintain energy homeostasis and the correct mitochondria number ^[5]. The balance between degradation and biogenesis of mitochondria is essential for healthy cells. On the contrary, when this balance falls, cells undergo snags. This is the case of a model of human stem cell differentiated RGCs (hRGCs) with the optineurin (a crucial actor for mitophagy) dominant mutation (E50K) ^[5]; this mutation was associated with normal-tension glaucoma ^[7]. In E50K hRGCs, restoring correct biogenesis through pharmacological treatment helps to balance energy homeostasis and counteract neurodegeneration ^[5]. In particular, in E50K hRGCs, inhibiting the tank-binding kinase 1 (TBK1) with the BX795 drug reduces cellular apoptosis, promotes mitochondrial biogenesis, increases mitochondrial mass, and minimizes mitochondrial swelling, which occurs when mitochondria increase their matrix volume to possibly produce more ATP. Also, in glaucomatous E50K hRGCs, activating the energy sensor AMPKα triggers, in turn, the mitochondrial biogenesis regulator PGC1α to maintain homeostasis. BX795 exerts its role independently from the AMPKα-PGC1α pathway. Interestingly, in non-mutant hRGCs in normal or under-stress conditions, BX795 treatment increased spare respiratory capacity and reduced apoptosis, suggesting an efficient neuroprotective strategy in non-genetic pathological conditions. Enhanced mitochondrial biogenesis was also observed in RGCs differentiated patient-specific human-induced pluripotent stem cells (hiPSCs) from MT-ND4-mutated Leber’s hereditary optic neuropathy (LHON)-affected patients ^[8]. The LHNO mutation reduces spare respiratory capacity, reducing mitochondria’s ability to supply energy and cell survival, promoting neurodegeneration. Furthermore, in this model, the expression of the antioxidant enzyme catalase appeared reduced, indicating higher oxidative stress and an imbalanced redox status. Additional studies have shown that MT-ND4 mutations induce elevated levels of oxidative stress that lead to dysfunctional and apoptotic RGCs ^[9]. Additionally, ROS inhibits the expression of the kinesin family member 5A (KIF5A) protein, causing an increment in the retrograde movement of the mitochondria in the axons. Also, the mix of the downregulation of KIF5A expression and the MT-ND4 mutation results in increased levels of apoptosis. The inhibition of the ERK1/2-Dynamin-related protein 1 (Drp1)–ROS axis was recently suggested as a potential therapeutic strategy to rescue RGC loss and counteract pathologically high intraocular pressure, a primary risk factor for glaucoma ^[10]. The detrimental effect of upregulation of p-ERK1/2 probably acts on Müller cells that, in turn, regulate the expression of p-Drp1 (Ser616) in RGCs. Drp1 is a mitochondrial protein that takes part in fission ^[11]. It was suggested that regulating its expression could help modulate mitochondrial ROS production and reduce RGC loss. Also, the Drp-1 protein was modulated via the A-Kinase anchoring protein 1 (AKAP1), a multifunctional mitochondrial scaffold protein that increases cell survival and regulates mitochondrial dynamics, bioenergetics, and mitophagy ^[12]. AKAP1 promotes mitochondrial elongation by regulating PKA/Drp1 anchoring, thus favoring mitochondrial good functioning. Furthermore, AKAP1 mediates mitochondrial bioenergetics by augmenting ATP synthesis and the mitochondrial membrane potential. Since oxidative stress and elevated intraocular pressure induce AKAP1 deficiency in RGCs and an increment of AKAP1 expression promotes RGC survival against oxidative stress, it was suggested that AKAP1 plays an essential role in mitochondrial preservation in RGCs during neurodegeneration induced by glaucoma. In RGCs, the loss of AKAP1 leads to an increment in Drp1 (Ser616) dephosphorylation, followed by mitochondrial fragmentation and loss ^[12][13]. On the contrary, phosphorylation of Drp1 mediated by AKAP1 promotes mitochondrial fusion and rescues RGCs from glaucoma insult ^[12]. In the glaucomatous DBA/2J retina and AKAP1^−/− mice, a significant increase in calcineurin (CaN) protein expression was detected, together with the dephosphorylation of Drp1 Ser637 ^[13]. In AKAP1^−/− mice, strong LC3 immunoreactivity was detected in RGC somas and axons, supported by the increment of LC3-II and the decrement of p62 levels, suggesting an enhancement in autophagosome production and, thus, the induction of autophagy/mitophagy. Furthermore, oxidative phosphorylation (OXPHOS) complexes (Cxs) deregulation was observed, with increased SOD2 protein expression, causing metabolic dysfunction and oxidative stress in the retina. Akt inactivation and Bim/Bax activation were also detected in AKAP1^−/− mice, contributing to glaucomatous neurodegeneration.

As well as in glaucoma neurodegeneration, RGCs are prone to deterioration in familial dysautonomia (FD), a disorder characterized by developmental and progressive neuropathies that affect the entire nervous system, also causing blindness ^[14]. FD is caused by a mutation in the IKBKAP/ELP1 gene, which encodes the inhibitor of κB kinase complex-associated protein IKAP, also named ELP1, involved in elongation and demanded the translation of codon-biased genes. In a mouse model of FD blindness, it was observed that the loss of IKAP caused progressive degeneration of RGCs with a progressive loss of mitochondrial membrane integrity, membrane potential, function, and thus ROS dysregulation. Interestingly, the other retinal neurons, including Müller glial, bipolar, amacrine, and photoreceptor cells, remained mainly uninjured, though with damaged mitochondria. This evidence supports that RGCs are highly vulnerable to mitochondrial dysfunction, probably due to their high energy demand and unique morphology. Recently, a case report described the thinning of the retinal nerve fiber and ganglion cell layers and decreased mitochondrial function in a 17-year-old patient ^[15]. This report associates the SIRT3 (sirtuin 3) gene mutation with mitochondrial optic neuropathy. SIRT3 is a well-known regulator of mitochondrial metabolism ^[16]. Supporting the concept that mitochondria biogenesis is crucial in maintaining homeostasis, it was observed that intravitreal transplanted iPSC-MSCs might donate functional mitochondria to RGCs in a mouse model of Leigh’s disease, contributing to protecting them from cell degeneration and death ^[17]. Furthermore, iPSC-MSCs reduce abnormal activation of Müller cells and inflammation by reducing neuroinflammatory cytokines such as TNF α, MIP−1g, GM-CSF, IL-5, IL-17, and IL-1 β, protecting RGCs from degeneration and loss. This evidence underlines a link between mitochondrial dysfunction, inflammation, and neurodegeneration. Glia neuroinflammation is a significant contributor to glaucoma. During elevated intraocular pressure conditions, an upregulation of TLR4 and IL-1β expression in Müller glia end feet was evident, both in the human glaucomatous retina and in the DBA/2J mouse that mimics human glaucoma ^[18]. At the same time, a significant decrement of apolipoprotein A-I binding protein (AIBP; gene name Apoa1bp) was detected in RGCs, leading to spatial vision dysfunction but not severe optic nerve damage. In Apoa1bp^−/− mice, AIBP deficiency activates mitochondrial fragmentation, mitochondrial cristae depletion, and energy production dysregulation, resulting in dysfunctional Müller glia and inflammatory conditions. AIBP deficiency impairs mitochondrial dynamics and decreases Cxs protein expression in the retina. In particular, in Apoa1bp^−/− RGC somas, mitochondrial fragmentation and reduced ATP production in RGCs were detected. Furthermore, an AIBP role was also suggested in the inner retina, affecting RGC dendrites in the IPL during glaucomatous neurodegeneration. In RGCs and the inner retina, AIBP deficiency contributes to oxidative stress, reducing SIRT3 and SOD2 amounts and increasing phospho-p38, a stress-signaling player, and ERK1/2. On the contrary, the administration of AIBP promotes RGC and inner retinal neuron survival and inhibits oxidative stress signaling and inflammatory responses in mice, which could result in neuroprotection. This evidence suggests that AIBP may have therapeutic prospects for treating glaucoma, blocking neuroinflammation, and acting on mitochondrial functions.

Correct mitochondrial functionality is crucial to preventing retinal neurodegeneration. It was demonstrated that counteracting mitochondrial fragmentation represents a chance to protect retinas from various malfunctions affecting cellular metabolism, cellular respiration, and apoptosis that participate in the development and progression of eye disease. In this context, each progress in molecular and functional investigations of RGCs represents a great opportunity for intervention against visual damage.